The work proposed for this grant focuses on the coordinate regulation of proliferation and apoptosis in rheumatoid arthritis (RA). Analysis of human RA synovial tissues (ST) sections revealed increased rates of synovial fibroblast proliferation and low rates of apoptosis, though the functional significance of reduced apoptosis remains to be elucidated. However, analysis of the rates of in vivo proliferation and apoptosis are limited in human-STs as tissue sections were taken late in disease course. Thus, utilization of animal models is vital for an understanding of the molecular pathways of proliferation and apoptosis in RA. Recently, adenoviral mediated delivery of Fas ligand (Ad-FasL), a known apoptotic inducer ameliorated experimental arthritis, suggesting that enhancing the rate of apoptosis by gene therapy may be a potential effective therapy. A caveat to Ad-FasL therapy is that high levels of Fas ligand is cytotoxic to many tissues of the body, thus development of other genes to be delivered to the RA joint is essential. We demonstrated that the anti-apoptotic protein and cell cycle modulator, Bcl-2 was highly expressed in RA compared with osteoarthritis synovial tissues, particularly in the CD68- negative, fibroblast-like synoviocyte population. In order to determine the importance of endogenous Bcl-2, an adenoviral vector expressing a hammerhead ribozyme to Bcl-2 (Ad-Rbz-Bcl-2) mRNA was employed. Ad-Rbz-Bcl-2 infection resulted in reduced Bcl-2 expression and cell viability in synovial fibroblasts isolated from RA-synovial tissues. In addition, Ad-Rbz-Bcl-2- induced mitochondrial permeability transition, cytochrome c release, activation of caspases 9 and 3, and DNA fragmentation. These data suggest that Bcl-2 is necessary for synovial fibroblast survival. In this proposal we describe studies to delineate the mechanism of the induction of mitochondrial permeabilty transition following Ad-Rbz-Bcl-2 infection. In addition we will investigate whether adenoviral mediated delivery of the Bcl-2 ribozyme is efficient in ameliorating adjuvant- induced arthritis in rats. The expected outcome is the suppression of AIA through the inhibition of fibroblast proliferation and increased apoptosis. This approach could lead to the development of a new therapeutic strategy for gene therapy in patients with rheumatoid arthritis.